Characterization of In Vitro Differentiation of Dental Papilla Cells In vitro techniques provide a well controlled environment for the study of cell differentiation. The odontoblasts are the cells responsible for dentin formation during normal development and during repair. The factor which induce the differentiation of these cells are not well understood. The long term goal is to elucidate the mechanism of odontoblast differentiation and identify factors that stimulate this process. These can then be used for clinical application for promotion of dentin repair. Dental papilla cells were removed from newborn rat molars and cultured on Type I collagen coated dishes containing DMEM supplemented with 10% FBS and antibiotics. Addition of ascorbic acid and dexamethasone resulted in the appearance of mineralized areas. These areas stained positive by Von Kossa and showed a high alkaline phosphatase activity. Xray microanalysis of the mineralized nodules revealed Ca an P as the prominent elements. Light and electron microscopic analysis reveal elongated cells with well developed ER and Golgi consistent with a well differentiated secretory cell. These observations suggest that dental papilla cells may differentiate into odontoblasts under these conditions. Biochemical, immunohistochemical and immunocytochemical identification of a dentin sialoprotein (a dentin specific marker protein) will be performed to confirm the odontoblast phenotype. The establishment of an in vitro system for examining odontablasts will allow the study of the effects of putative pulp treatment agents on normal odontoblast differentiation and dentin formation. Keys words: odontoblast, dental papilla, tissue culture, electron microscopy